5.-CONCLUSIONS
The results presented in this work have evidenced the beneficious effect
of antiapoptotic gene BHRF1 in hybridoma cultures in terms of cell
growth, mAb productivity and metabolism; characterized by a more
efficient usage of nutrients, doubling the cell density in culture under
identical conditions and same culture media. The specific productivity
of the cells has been multiplied by two, obtaining three times increase
of final IgG3 concentration.
The study and comparison of the intracellular fluxes by means of the
Flux Balance Analysis of the KB26.5 and KB26.5-BHRF1 engineered cell
lines has enlightened the interactions and effects of BHRF1 protein on
the metabolic pathways. In short, BHRF1 primarily affected glucose
uptake rate, reducing glycolysis by 50% and consequently reducing the
lactate generation by more than 60%. Interestingly, the total ATP
generation in the engineered KB26.5-BHRF1 cell line has been
significantly decreased due to the lower energy requirements for
maintenance, probably due to the lower energy requirements for
maintaining ion gradients, although this hypothesis should be evaluated
with further metabolic flux analysis experiments.
The use of antiapoptotic genes has been extensively explored in the last
years, as manifested in a very recent review about the attenuation of
apoptosis in CHO cell line (Henry et al., 2020). Many efforts have been
displayed to understand how these genes affect cell apoptosis; but the
impact of those in protein production and specially in metabolism have
not been completely understood by the scientific community. To tackle
this issue, a strategy to obtain the isogenic cell lines should be
implemented as it would allow the comparison between different
anti-apoptotic genes and their effect.